Department of Molecular Biology and Biochemistry, University of California, Irvine, California, USA
mBio. 2011 Feb 22;2(1):e00341-10. doi: 10.1128/mBio.00341-10. Print 2011.
Murine leukemia viruses (MuLVs) encode two forms of Gag polyprotein: the precursor for the viral core proteins (Pr65(gag) for Moloney MuLV [M-MuLV]) and a longer glycosylated form (glyco-gag, or gPr80(gag)). gPr80(gag) is translated from the same unspliced viral RNA as Pr65(gag), from an upstream in-frame CUG initiation codon. As a result, gPr80(gag) contains 88 unique N-terminal amino acids that include a signal peptide that conducts gPr80(gag) into the rough endoplasmic reticulum, where it is glycosylated, exported to the cell surface, and cleaved into two proteins of 55 and 40 kDa. The amino-terminal 55-kDa protein remains cell associated with the 88 unique amino acids exposed to the cytosol. We previously showed that gPr80(gag) facilitates efficient M-MuLV release through lipid rafts. In this report, we found that the unique N-terminal domain of gPr80(gag) is sufficient to facilitate enhanced M-MuLV particle release from transfected 293T cells. A search for cellular proteins involved in gPr80(gag) function led to cellular La protein. Overexpression of mouse or human La enhanced M-MuLV particle release in the absence of glyco-gag, and the released virus had a reduced buoyant density characteristic of increased cholesterol content. Moreover, small interfering RNA (siRNA) knockdown of human La abolished glyco-gag enhancement of M-MuLV release. These results implicate La as a cellular protein involved in M-MuLV glyco-gag function. We also found that overexpression of mouse or human La could enhance HIV-1 release in the absence of gPr80(gag). Therefore, M-MuLV and HIV-1 may share a pathway for release through lipid rafts involving La.
鼠白血病病毒(MuLVs)编码两种形式的 Gag 多聚蛋白:病毒核心蛋白的前体(Pr65(gag),用于 Moloney MuLV [M-MuLV])和一种更长的糖基化形式(glyco-gag,或 gPr80(gag))。gPr80(gag)与 Pr65(gag) 从相同的未剪接病毒 RNA 翻译而来,来自上游的一个框内 CUG 起始密码子。因此,gPr80(gag)包含 88 个独特的 N 端氨基酸,其中包括一个信号肽,该信号肽将 gPr80(gag)引导至粗面内质网,在那里进行糖基化,然后输出到细胞表面,并切割成 55 和 40 kDa 的两种蛋白质。N 端 55 kDa 的蛋白质与暴露在细胞质中的 88 个独特氨基酸保持细胞相关。我们之前曾表明,gPr80(gag)通过脂筏促进 M-MuLV 的有效释放。在本报告中,我们发现 gPr80(gag)的独特 N 端结构域足以促进转染 293T 细胞中 M-MuLV 颗粒的增强释放。寻找参与 gPr80(gag)功能的细胞蛋白导致发现细胞 La 蛋白。在没有糖基化 gPr80(gag)的情况下,过量表达小鼠或人 La 增强了 M-MuLV 颗粒的释放,并且释放的病毒具有特征性的较低浮力密度,表明胆固醇含量增加。此外,人 La 的小干扰 RNA (siRNA) 敲低消除了糖基化 gPr80(gag)增强 M-MuLV 释放的作用。这些结果表明 La 是参与 M-MuLV 糖基化 gPr80(gag)功能的一种细胞蛋白。我们还发现,过量表达小鼠或人 La 可以在没有 gPr80(gag)的情况下增强 HIV-1 的释放。因此,M-MuLV 和 HIV-1 可能通过涉及 La 的脂筏共享一种释放途径。